microrna mir 149  (Vector Biolabs)

Journal: PLoS ONE

Article Title: XB130, a New Adaptor Protein, Regulates Expression of Tumor Suppressive MicroRNAs in Cancer Cells

doi: 10.1371/journal.pone.0059057

Figure Lengend Snippet: Significantly upregulated miRNAs in XB130 shRNA transfected Cells.

Article Snippet: As for quantifications of the mature miRNAs, a TaqMan® MicroRNA RT Kit and TaqMan® MicroRNA assays specific for hsa-miR-33a, has-miR-149 and has-miR-193a-3p (Applied Biosystems) were used for RT reactions.

Techniques: shRNA, Transfection

Journal: PLoS ONE

Article Title: XB130, a New Adaptor Protein, Regulates Expression of Tumor Suppressive MicroRNAs in Cancer Cells

doi: 10.1371/journal.pone.0059057

Figure Lengend Snippet: The mature forms ( A ) and primary transcripts ( B ) of miR-33a, miR-149 and miR-193a-3p are significantly higher in XB130 shRNA transfected WRO cells (C3-1, C3-4, C4-3 and C4-11) than in WRO (*P<0.05).

Article Snippet: As for quantifications of the mature miRNAs, a TaqMan® MicroRNA RT Kit and TaqMan® MicroRNA assays specific for hsa-miR-33a, has-miR-149 and has-miR-193a-3p (Applied Biosystems) were used for RT reactions.

Techniques: shRNA, Transfection

Journal: PLoS ONE

Article Title: XB130, a New Adaptor Protein, Regulates Expression of Tumor Suppressive MicroRNAs in Cancer Cells

doi: 10.1371/journal.pone.0059057

Figure Lengend Snippet: MRO cells were transfected with GFP vector alone, or GFP-XB130, GFP-XB130ΔN (XB130 N-terminus deletion mutant). GFP positive cells were collected by FACS. ( A ) Western blotting revealed that MRO cells express very low XB130. Transfection of GFP-XB130 (MRO-XB130) or GFP-XB130ΔN (MRO-XB130ΔN) showed XB130 expression in MRO cells. The mature forms ( B ) and primary transcripts ( C ) of miR-33a, miR-149 and miR-193a-3p were examined by real-time quantitative RT-PCR. These miRNAs and pri-miRNAs in MRO cells were significantly higher than that in WRO cells, and significantly reduced by overexpression of GFP-XB130, but not by GFP-XB130ΔN. *P<0.05 compared with WRO cells.

Article Snippet: As for quantifications of the mature miRNAs, a TaqMan® MicroRNA RT Kit and TaqMan® MicroRNA assays specific for hsa-miR-33a, has-miR-149 and has-miR-193a-3p (Applied Biosystems) were used for RT reactions.

Techniques: Transfection, Plasmid Preparation, Mutagenesis, Western Blot, Expressing, Quantitative RT-PCR, Over Expression

Journal: PLoS ONE

Article Title: XB130, a New Adaptor Protein, Regulates Expression of Tumor Suppressive MicroRNAs in Cancer Cells

doi: 10.1371/journal.pone.0059057

Figure Lengend Snippet: ( A ) Transfection of miR mimics (33a-m, 149-m and 193a-m) significantly increased respective miRNA expression, whereas control miR mimic (cont-m) had no such effect, as quantified by real-time quantitative RT-PCR. ( B ) Western blotting shows that transfection of miR mimic did not change XB130 protein levels. ( C–E ) MYC, FOSL1 and SCL7A5 are predicted target of miR-33a, miR-149 and miR-193-3p, respectively. Upper panel shows examples that the expression of the predicted target protein was decreased by respective miR mimic transfections as determined by Western blotting. Lower panel shows the quantification of the expression levels by densitometry.

Article Snippet: As for quantifications of the mature miRNAs, a TaqMan® MicroRNA RT Kit and TaqMan® MicroRNA assays specific for hsa-miR-33a, has-miR-149 and has-miR-193a-3p (Applied Biosystems) were used for RT reactions.

Techniques: Transfection, Expressing, Quantitative RT-PCR, Western Blot

Journal: PLoS ONE

Article Title: XB130, a New Adaptor Protein, Regulates Expression of Tumor Suppressive MicroRNAs in Cancer Cells

doi: 10.1371/journal.pone.0059057

Figure Lengend Snippet: ( A ) Seed sequences of miR-33a, miR-149 and miR-193a-3p and pairing 3′UTRs sequence of MYC, FOSL1 and SLC7A5 as predicted by TargetScan are shown. The pmirGLO construct with the cloned 3′UTR sequences (luc-MYC, luc-FOSL1 and luc-SLC7A5) and the miR mimic (33a-m, 149-m and 193a-m) or control miR mimic (cont-m) were co-transfected into WRO cells. ( B ) Overexpression of miR-33a significantly reduced luciferase activity of the pmirGLO construct with cloned 3′UTR sequences of MYC (luc-MYC). Similarly, overexpression of miR-149 and miR-193-3p significantly reduced luciferase activity of the luc-FOSL1 ( C ) and the luc-SCL7A5 ( D ), respectively.

Article Snippet: As for quantifications of the mature miRNAs, a TaqMan® MicroRNA RT Kit and TaqMan® MicroRNA assays specific for hsa-miR-33a, has-miR-149 and has-miR-193a-3p (Applied Biosystems) were used for RT reactions.

Techniques: Sequencing, Construct, Clone Assay, Transfection, Over Expression, Luciferase, Activity Assay

Journal: PLoS ONE

Article Title: XB130, a New Adaptor Protein, Regulates Expression of Tumor Suppressive MicroRNAs in Cancer Cells

doi: 10.1371/journal.pone.0059057

Figure Lengend Snippet: The numbers of viable cells were assessed by MTS assay (A) and cell counting (B) at 24, 48, and 72 h post-transfection of control mimic (cont-m) and specific miR mimic (33a-m, 149-m and 193a-m). The overexpression of miR-33a, miR-149 and miR-193a-3p led to reduction in cell proliferation. ( C ) Cell proliferation markers, Ki-67 and PCNA levels, were also reduced in miR mimic treated cells (33a-m, 149-m and 193a-3p). ( D ) Apoptosis was determined by flow cytometry using PI/annexin V double staining. Overexpression of miR-193a-3a significantly induced both early apoptosis (annexin V positive/PI negative) and late apoptosis (annexin V/PI double positive) in WRO cells. *: P<0.05.

Article Snippet: As for quantifications of the mature miRNAs, a TaqMan® MicroRNA RT Kit and TaqMan® MicroRNA assays specific for hsa-miR-33a, has-miR-149 and has-miR-193a-3p (Applied Biosystems) were used for RT reactions.

Techniques: MTS Assay, Cell Counting, Transfection, Over Expression, Flow Cytometry, Double Staining